! PROJECT DESCRIPTION Obesity is a chronic organismal stress that impacts multiple biological functions and tissues. Obesity and its sequelae modulate the immune system and the hematopoietic activity in the bone marrow (BM). Notably, obesity has been associated with altered immunological functions and an overall increased risk for hematological malignancies. However, despite their clinical relevance, the mechanisms by which obesity affects the health of the hematopoietic system and might contribute to its pathological dysregulation have yet to be characterized. Here, we seek to decipher the impact of obesity on the molecular and cellular fitness of the hematopoietic stem cell (HSC) compartment. In Aim1, we will investigate the cell-intrinsic molecular mechanisms that allow HSCs to cope with the aberrant environmental stresses triggered by obesity. Based on our preliminary results, we will focus on the transcriptional factor Gfi1 as we found that up-regulation of this factor is crucial in regulating HSC fate in obesity. Notably, we uncovered a novel molecular circuit that links oxidative stress and Gfi1 expression in HSCs. Here we will determine the molecular mechanisms underlying this link in obesity. We will also test the hypothesis that Gfi1 up-regulation affects the long-term fitness of the HSCs by modulating their ability to mount proper stress responses. In Aim2, we will investigate at the single cell level how obesity disrupts the homeostasis of the HSC compartment. We will determine whether the obese environment alters the molecular and functional, clonal diversity of the HSC compartment, leading to the development of a clonal hematopoiesis. Finally, we will test the hypothesis that the obese environment could favor the emergence of clones carrying pre-neoplastic features. Altogether, our studies will elucidate the impact of obesity on the hematological system. They will test the hypothesis that obesity alters the HSC intrinsic regulatory programs, affects the clonal structure of the HSC pool and promotes the emergence of pre- leukemic HSC clones. These studies stand to make critical contributions to our understanding of the adverse effects of chronic obesity on the health of the HSC compartment. In the midst of a global obesity epidemic, the long-term goal of this project is to decipher the potential risks associated with the use of obese donors for BM stem cell transplantation and the influence of obesity on the long-term development of pre-neoplastic hematopoiesis.